Unprocessed insulin proreceptors due to point mutation at the cleavage site

Two sisters presented with severe insulin resistance and markedly decreased insulin binding to erythrocytes, cultured fibroblasts and transformed lymphocytes. The dose-response curve of insulin-stimulated amino acid uptake in the fibroblasts was shifted to the right. The molecular weight of the insulin receptor on the transformed lymphocytes from the patients was 210,000 and could not be dissociated to alpha- and beta-subunits by dithiothreitol treatment. However, the proreceptor was cleaved by trypsin and this led to the production of alpha-subunit with normal insulin binding. We performed cDNA sequence analysis of the cleavage site of the insulin proreceptor from the patients. The polymerase chain reaction was used to obtain a large amount of cDNA coding for the region including the interconnecting site. A thermostable DNA polymerase, Taq polymerase, successfully produced enough cDNA for the region to be sequenced. The results showed an AGG (Arg) to AGT (Ser) point mutation, resulting in the change of the interconnecting sequence of the two subunits from -Arg-Lys-Arg-Arg- to -Arg-Lys-Arg-Ser-. These results suggest that the tertiary structure change of the cleavage site leads to production of unprocessed insulin proreceptors.